TY - THES A1 - Ratnayake, Hasiru T1 - A Systematic Review of Rapamycin- Mediated Fetal Hemoglobin Induction via the mTOR Pathway N2 - mTOR inhibitor rapamycin has been observed to induce fetal hemoglobin in patients of β-hemoglobinopathies, a group of disorders in which mutations in the gene responsible β-globin chains results in reduced or malformed RBC production, leading to anemia. The induction of fetal hemoglobin bypasses these mutations, as they are found on the gene responsible for adult hemoglobin. Following birth, the fetal hemoglobin gene is silenced, and the adult gene is activated through a process known as the hemoglobin switch. Rapamycin treatment reverses this hemoglobin switch but the mechanism by which it does so is currently unknown. Rapamycin functions through its inhibition of mTORC1, a protein complex responsible for nutrient- and growth factor-dependent cell growth and proliferation. This literature review seeks to collate existing information on the mTORC1 signalling pathway and the hemoglobin switch, allowing for a specific mechanism by which rapamycin induces fetal hemoglobin through its inhibition of mTORC1 to be hypothesized. Following the literature search, 55 studies on the mTORC1 pathway and 31 studies on the hemoglobin switch were summarized. Using the information gained from this, three hypotheses were put forward for the possible induction mechanism, out of which the primary hypothesis is based on the hemoglobin switching factor KLF1. This was based on the observation that the knockdown of Raptor, a crucial component of mTORC1, in mice caused a decreased expression of KLF1. As KLF1 is an essential component of the hemoglobin switch, this provides a direct link between the inhibition of mTORC1 and the reversal of the hemoglobin switch. Additionally, the efficiency of rapamycin in comparison to other methods of fetal hemoglobin was discussed, including newer generations of mTOR inhibitors developed using rapamycin as a foundation. Finally, possible avenues of research to confirm the validity of the proposed hypotheses and to address points of ignorance within each topic were discussed. Y1 - 2024 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:hbz:1383-opus4-21155 ER - TY - THES A1 - Mumtaz, Muhammad Imran T1 - Generation and expression of a mutated murine tRNA-guanine transglycosylase variant N2 - Translation of messenger RNA carrying the nucleic code for protein synthesis is one of the vital bioprocesses of living organisms. This translation is mediated by transfer RNA, which allocates specific amino acids to the growing chain by decoding the genetic code. The tRNAs with the G34U35N36 anticodon sequence are recognised by an enzyme called tRNA-guanine transglycosylase, which catalyses the exchange of guanine with queuine. TGT enzyme is found in all three domains of life and is involved in post-transcriptional modification. TGT is of keen interest in the development of therapeutics against shigellosis, since the genetic inactivation of TGT leads to decreased pathogenicity in Shigella. For understanding the differences between bacterial and eukaryotic TGT, studies have been performed in the past, which show remarkable resemblance. On the contrary, some pronounced structural contrast can also be observed. One prominent difference is that the bacterial TGT contains a short turn of three amino acids, whereas, in eukaryotes, this turn is replaced by an extended loop of more than 45 amino acids. The function of this loop in eukaryotic TGT is not known. The loop is a highly flexible region and, thus, is not structurally solved. To elucidate the function of this loop, a TGT mutant lacking this loop was generated, expressed and purified. TGT mutant was expressed in different bacterial cell strains. For the purification, different chromatography techniques were utilised. The mutation resulted in a low yield of the protein in comparison to the wild-type TGT. This leads to the inference that the loop influences the physicochemical characteristics of the protein. KW - RNA-guanine transglycosylase KW - Generation KW - Expression KW - Queuine KW - Protein purification KW - Chromatography KW - Site-directed mutagenesis KW - Protein mutation Y1 - 2023 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:hbz:1383-opus4-18560 ER - TY - THES A1 - May, Clarissa T1 - Efficacy of Traditional Chinese Medical DrugsThe Treatment of Common Illnesses in Comparison with Western medicine N2 - The efficacy of treating common illnesses with traditional Chinese medical drugs has been a subject of growing interest and debate especially when compared to established practices of modern Western medicine. The integration of alternative and traditional medicine practices also gains significance as healthcare systems evolve. This literature review aims to comprehensively assess and compare the effectiveness of traditional Chinese medical drugs with modern Western medial drugs in addressing common illnesses. Through a systematic analysis of reviewed scientific articles, clinical studies, and relevant literature the effectiveness of traditional Chinese medical drugs will be identified and compared to modern Western medical drugs and treatment options in context of the treatment success rates, symptom relief, adverse effects, and patient satisfaction. The study focuses on the herbal based remedies and their traditional therapeutic treatment methods, to illuminate their mechanisms of action and potential therapeutic benefits for a range of common illnesses such as the common cold and flu, digestive disorders, musculoskeletal pain, and skin conditions. Furthermore, the potential synergies between the two practices will be explored, as integrative healthcare becomes increasingly important as a comprehensive approach to patient wellbeing. Through a comprehensive analysis of the available medical literature and clinical data, this paper attempts to address the potential of traditional Chinese medical drugs as an alternative or complementary method to modern Western medicine for various common illnesses. The findings may contribute to informed decision-making by healthcare professionals and patients, offering insights into the strengths and limitations of each approach. Additionally, the research aims to foster a deeper understanding of cross-cultural medical practices and their potential integration into contemporary healthcare systems. KW - Traditional Chinese medicine Y1 - 2024 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:hbz:1383-opus4-19440 ER - TY - THES A1 - Alshaikh, Badran-Mohammednoor T1 - Medi: A Software System for Improving the Analysis and Interpretation of Medical Tests in Clinical Laboratories. N2 - The rapid increase in patient numbers and the complexity of clinical information, coupled with a significant lack of assistive technologies for routine procedures, has emphasized the urgent need for innovative technological solutions in clinical laboratories. This need is amplified by the existence of over 238 blood tests, highlighting the challenge of managing and interpreting vast amounts of clinical information and data efficiently. The primary objective of this research is to evaluate the effectiveness of Medi, a state-of-the-art software system designed to enhance the workflow of medical routine tasks in the pre-testing steps in clinical laboratories. By focusing on routine hematology, biochemistry, and microbiology tests, the study aims to revolutionize clinical laboratory processes, improve diagnostic accuracy, and enhance patient care. Through an extensive approach regarding specimen collection of blood and the utilization of automated medical devices, this study sheds light on the advantage of Medi to streamline operations within clinical laboratories. The research methodically assesses the impact of Medi on laboratory workflows and patient outcomes, employing a user-centric design to ensure the software meets the practical needs of healthcare professionals. Medi demonstrated significant improvements in laboratory efficiency, with streamlined workflow. Key findings include the standardization of blood specimen collection procedures, limiting blood-tube-selection errors, and a positive impact on laboratory operations. The software facilitated a more cohesive integration of clinical data, contributing to better-informed decision-making and improved patient care. By comparing these outcomes with the current state of clinical diagnostics, Medi's introduction represents a transformative shift in laboratory operations. It not only addresses the pressing need for advanced assistive technologies in the clinical laboratory settings but also sets a new standard in patient care. This research underlines the potential of integrating software solutions with existing laboratory technologies, paving the way for a new era in clinical testing. Y1 - 2024 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:hbz:1383-opus4-19466 ER - TY - THES A1 - Schiffers, Lukas T1 - Optimization of RT-PCR Assays for the Roche cobas® 6800/8800 Utility Channel: Influence of 2´-O-Methyl modified Primers on PCR efficiency N2 - Automated, high-throughput real-time PCR instruments such as the cobas® 6800/8800 from Roche have revolutionized the operation in molecular diagnostics laboratories as they provide a complete process of patient samples including all necessary steps for PCR applications from material preparation up to the calculation and evaluation of final results with a minimum of user interaction. In addition to the broad spectrum of commercially available ready-to-use reagent kits for blood screenings, virologic and microbiological assays as well as tests designed for women’s health, the cobas® 6800/8800 provides a tool which allows users to develop and implement their own tests onto the high-throughput device. Equipped with this tool, called the cobas® omni Utility Channel, these automated instruments serve a high flexibility and can be adjusted to users’ personal needs. Notwithstanding the advantages of the Utility Channel, a limited number of lab developed tests have been implemented onto the cobas® 6800/8800 so far as no guideline for the optimization and transfer of such tests onto the Utility Channel has been provided to the public yet. In the course of this bachelor thesis, four real-time PCR assays, including an allele-specific duplex test for HLA-B27, an established in-house test for the differentiation and detection of Ureaplasma urealyticum/parvum, a published assay for the detection of Chlamydia trachomatis as well as a duplex reverse-transcription assay for the detection and differentiation of norovirus genogroup one and two have been optimized for the use on Utility Channel. Furthermore, 2´-O-Methyl modified primers have been integrated into the optimization process of all assays to asses their influence on PCR efficiency. Three of these four assays have been transferred successfully onto a cobas® 6800 following a developed method which lead to the final design of a flowchart guideline for the optimization of lab developed tests for the use on a cobas® 6800/8800 system. KW - PCR efficiency KW - Modified primers KW - Methylated primers KW - Cobas KW - Molecular biology Y1 - 2020 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:hbz:1383-opus4-4632 ER - TY - THES A1 - Khedher, Lara T1 - Evolution of CRISPR-Cas genome editing methods: Chances and challenges in biotechnology N2 - Clustered regularly interspaced short palindromic repeats, CRISPR, are located on DNA sequences, which encode for an antiviral defense mechanism in bacteria and archaea. Together with CRISPR-associated proteins, Cas endonucleases, CRISPR-Cas has emerged into a powerful platform for editing DNA and RNA. This thesis investigates the evolution and function of this prokaryotic defense mechanism and points out the three most applied CRISPR-Cas systems in relation to current applications in biotechnology. Furthermore, chances and challenges of named systems as well as a guideline for choosing the appropriate system with Cas endonucleases is framed. Therefore, several scientific databases were screened for the most informative and influential publications around the topic. Information was collected and organized in a reference management software. The utilized literature was tabularly denoted and gives an overview of the broad scope of CRISPR-Cas. The analysis of the topic shows, that CRISPR-Cas offers a broad application range with a huge chance for genome editing. A conclusion of this thesis is, that CRISPR-Cas has an enormous potential to be implemented for human genome editing. However, its challenges should be cleared beforehand. Further research has to be done to fully understand the impact CRISPR-Cas may have in future. KW - Genome editing KW - Genetics KW - CRISPR KW - Cas Y1 - 2020 ER - TY - THES A1 - Ranganathan, Shraddha T1 - Evaluation of two methods to reduce legume-related flatulence through enzymatic digestion of flatulence factors N2 - Legumes are nutritionally equivalent to many meat products, and can be used to supplement or replace meat in daily diets. This is beneficial for a variety of reasons, such as: i) growing legumes can help to reduce acidification of soil, global warming potential and energy use; ii) livestock is taxing on the environment in terms of adding to the volume of greenhouse gases and nitrification of soil while legume crops fix soil nitrogen. Consumers can be opposed to adding legumes to their diet due to the perception of legumes causing flatulence. Intestinal gas buildup, bloating, cramps, abdominal pain and flatulence are caused by raffinose family oligosaccharides (RFOs) which cannot be digested in monogastric organisms such as humans. They are therefore broken down by microflora in the intestine; this bacterial digestion releases large volumes of hydrogen, which causes flatulence. The human body lacks the enzyme required to break down these RFOs — ⍺-galactosidase. This experiment evaluated two methods of applying ⍺-galactosidase to RFOs before they reach the intestinal microflora. The first method evaluates the effectiveness of enzymatically digesting the legumes before consumption. The second evaluates the effectiveness of the enzyme supplement Beano, which applies the enzyme to RFOs in the stomach. Experimental data showed that enzymatically digesting raw flours does significantly reduce the amount of RFOs in the legume. Similarly, Beano also reduces RFOs significantly. In 4 out of the 6 legumes sampled, there was no significant difference between the two methods. In order to consider the methods for commercial use, other factors (such as economic, logistical, etc.) must also be considered. At the outset, it appears that taking an enzyme supplement such as Beano might be more economically viable in the long term for the consumer, since processing costs for the flatulence free legumes would drive up the price of (normally cheap) legumes. There is an increase in the amount of people who are giving up meat for environmental and other reasons. For these consumers, as well as those who come from cultures that integrate legumes in their cuisine, the removal of flatulence factors from this nutrition-rich food group will be very beneficial. KW - Alpha-galactosidase KW - Bloating KW - Enzymatic assay Y1 - 2020 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:hbz:1383-opus4-4903 ER - TY - THES A1 - Karssli, Mahmoud T1 - CRISPR technology in human medicine: advantages and disadvantages over traditional treatments N2 - Genome editing tools hold tremendous potential in fighting various diseases via editing specific sites within the target gene of interest of distinct cell types. Among these tools, CRISPR has shown several advantages over alternative nucleases, such as ease of design, high efficiency, and low cost. In this work, a comprehensive review of the existing research on the application of CRISPR technology to the treatment of human diseases is presented. These diseases include monogenic and infectious diseases, as well as conditions for which immunotherapies are typically applied, such as cancer and autoimmune disorders. Following this review is an analysis of the advantages and disadvantages of CRISPR-based strategies compared to traditional treatments in a selection of diseases; this aims to demonstrate the overall potential of CRISPR therapeutic applications in humans and the need for further investigation and development. Y1 - 2022 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:hbz:1383-opus4-16080 ER - TY - THES A1 - Kanwal, Hafsa T1 - Detection of the Fish Pathogen Vagococcus salmoninarum via Quantitative real-time PCR for Aquaculture N2 - PHOTO-SENS is a project that aims to develop photonic biosensing chips as a means of allowing onset detection of pathogenic infections in aquaculture. These chips are meant to be a way for quick and simple routine testing that would enable aquaculture facilities to recognize potential outbreaks and minimize their effects. The chips detect target bacteria using short DNA sequences known as probes which are complementary to target DNA. One of the fish pathogens under consideration for the course of this project is Vagococcus salmoninarum, that is a causative agent of Vagococcosis. For this work, a primer and probe set specific to V. salmoninarum was designed. These primers and probes were then tested via qPCR for their selectivity and sensitivity and most importantly, quantification of DNA in samples using two different standard dilution series. Due to V. salmoninarum being a Gram-positive bacteria, three methods of DNA extraction were used and compared for maximum efficiency. Moreover, experiments were also conducted to mimic capture of eDNA (environmental DNA) from highly diluted water samples. Upon comparing the three DNA extraction methods, the improved 5% Chelex protocol that included rapid agitation via glass beads, yielded the highest amount of DNA that was closest to expected SQ. The detection of all samples, including very low copy number, was possible via qPCR. The first standard series which was the gBlocks standard series had the lowest detection limit of 1 copy with an efficiency of 105.8% and the second standard series that is the CFU standard series had a detection limit of 10 copies and had an efficiency of 104.2%. The filtration experiment also showed detection in diluted, filtered samples. Since detection using the designed probe was successful, the probes are therefore considered suitable for detection of Vagococcus salmoninarum in the PHOTO-SENS project. Y1 - 2022 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:hbz:1383-opus4-16222 ER - TY - THES A1 - Lavasanifar, Parsa T1 - Comparative analysis of guide RNA design tools N2 - Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR), has rapidly come to the forefront of genome engineering and has revolutionized the field. Today, CRISPR-Cas is regarded to be the most effective and efficient tool in genome editing. It is considered to have many applications in medicine, agriculture and biotechnology. The Cas protein and gRNA are two of the most crucial components of the CRISPR-Cas system. Currently, there are barriers that prevent the full potential of CRISPR-Cas systems in clinical applications, including, ethical concerns, off-target effects, immune system response, method of delivery. Off-target prevention relies heavily on gRNA analysis. With the help of technology and algorithms developed based on researches, there have been many bioinformatical tools focused on the off-target activity inhibition and increasing efficiency. In order to achieve practical use of CRISPR systems, there needs to be more predictive bioinformatical tools to minimize the off-target activity. This thesis aims to review some of these tools and discusses the criteria for future bioinformatical tools’ development by addressing the current limitations and challenges. This thesis is written theoretically by using various reputable resources such as PubMed in English. Because many of the tools covered in this thesis had no prior reviews, the author had to learn how to use them, therefore this thesis does not cover all of their features for professional users. In the result section, the author has provided figures from the interface of the tools along with description of the functionality of the algorithms in use. All of the mentioned tools are then examined in the discussion part. There, features such as speed, ease of use for beginners, price, being open-source, being web or local based, flexibility in working with other tools and batch analysis are reviewed and compared. There short conclusive results are also depicted in a table for effective comparison. Y1 - 2022 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:hbz:1383-opus4-15307 ER -